Bromophenol coupled with diketopiperazine from marine red alga symphyocladia latiuscula

Article abstract of DOI:10.1007/s10600-012-0327-9

2-(2,3,6-Tribromo-4,5-dihydroxybenzyl)hexahydropyrrolo[1,2-a]pyrazine-1, 4-dione (1), a new bromophenol C-N coupled with diketopiperazine, was isolated from the ethanol extract of marine red alga Symphyocladia latiuscula. Its structure was elucidated by s

Full text of DOI:10.1007/s10600-012-0327-9

Chemistry of Natural Compounds, Vol. 48, No. 4, September, 2012 [Russian original No. 4, July–August, 2012]
BROMOPHENOL COUPLED WITH DIKETOPIPERAZINE
FROM MARINE RED ALGA Symphyocladia latiuscula
1*
1
1
Xiuli Xu, Liyuan Yin, Nianqiao Fang,
UDC 547.565.2
3
2*
Xiao Fan, and Fuhang Song
2-(2,3,6-Tribromo-4,5-dihydroxybenzyl)hexahydropyrrolo[1,2-a]pyrazine-1,4-dione (1), a new bromophenol
C-N coupled with diketopiperazine, was isolated from the ethanol extract of marine red alga Symphyocladia
latiuscula. Its structure was elucidated by spectroscopic analysis, including high-resolution mass spectroscopy
and 1 and 2-dimensional NMR techniques. The absolute configuration of 1 was assigned using Marfeyꢀs
method on its acid hydrolysate.
Keywords: Symphyocladia latiuscula, bromophenol, diketopiperazine.
The marine red alga Symphyocladia latiuscula is a rich source of bromophenols of several structural types and with
various biological activities. Previous chemical studies on this species have resulted in the characterization of 15 monoaryl
and diaryl bromophenols [1–7]. Some of them showed aldose reductase inhibitory activity [1], antibacterial activity [2, 7], and
free-radical-scavenging activity [3–6]. As part of our recent program to assess systematically the chemical and biological
diversity of seaweeds distributed along the gulf of the Yellow Sea, one new bromophenol has been isolated from Symphyocladia
latiuscula [7]. Continuing our investigation of the same material, we report herein the isolation and structural elucidation of a
new bromophenol C-N coupled with diketopiperazine (1). This is the first report of bromophenols coupled with diketopiperazine
derivatives.
Br
3'
5
OH
OH
Br
3
O
7 N
5'
1'
7'
1N
O
Br
9
1
Compound 1 was obtained as a brown amorphous powder. The positive ESI-MS spectrum exhibited a characteristic
+
tribrominated quasi-molecular ion peak cluster at m/z 511/513/515/517 (1:3:3:1) [M + H] , and the molecular formula was
79
+
determined as C H
Br N O by positive HR-ESI-MS at m/z 510.8509 for [M + H] . The IR spectrum showed an absorption
14 13
3 2 4
–1
–1
band for hydroxyl groups at 3247 cm , the characteristic absorption bands for an aromatic ring at 1554 and 1450 cm , as
well as the absorption bands for the carbonyl group at 1660 and 1718 cm . The H NMR spectrum of 1 in DMSO-d showed
–1
1
6
two typical AB spin coupling systems attributed to two isolated methylenes at ꢁ 5.05 (1H, d, J = 15.0 Hz) and 4.71 (1H, d,
J = 15.0 Hz) for H -7ꢀ, at ꢁ 3.92(1H, d, J = 15.0 Hz) and 3.17 (1H, d, J = 15.0 Hz) for H -9, three methylenes at ꢁ 2.22 (1H, m) and
2
2
1.91 (1H, m) for H -4, 1.84 (2H, m) for H -5, 3.39 (1H, m) and 3.31 (1H, m) for H -6, as well as a methine at ꢁ 4.20 (1H,
2
2
2
t, J = 7.0 Hz). The protonated carbons and their corresponding protons were unambiguously assigned by the HSQC experiment.
1) School of Ocean Sciences, China University of Geosciences, 100083, Beijing, P. R. China, fax: +86 10 82320065,
e-mail: xuxl@cugb.edu.cn; 2) Institute of Microbiology, Chinese Academy of Sciences, 100101, Beijing, P. R. China,
fax: +86 10 62538564, e-mail: songfuhang@im.ac.cn; 3) Institute of Oceanology, Chinese Academy of Sciences, 266071,
Qingdao, P. R. China. Published in Khimiya Prirodnykh Soedinenii, No. 4, July–August, 2012, pp. 560–561. Original article
submitted June 20, 2011.
622
0009-3130/12/4804-0622 02012 Springer Science+Business Media, Inc.

1
13
TABLE 1. H and C NMR Spectral Data of Compound 1, J/Hz*
C atom
HMBC
C atom
HMBC
ꢁC
ꢁH
ꢁC
ꢁH
2
3
4
167.0 s
58.3 d
28.4 t
124.3 s
117.5 s
114.4 s
144.2 s
146.9 s
114.2 s
50.1 t
1ꢀ
2ꢀ
4.20 (1H, t, J = 7.0)
2.22 (1H, m)
1.91 (1H, m)
1.84 (2H, m)
3.39 (1H, m)
3.31 (1H, m)
4, 7
5,
2, 5
ꢀ
ꢀ
ꢀ
3
4
5
5
6
22.2 t
44.6 t
6ꢀ
7ꢀ
5.05 (1H, d, J = 15.0)
4.71 (1H, d, J = 15.0)
ꢀ
ꢀ
ꢀ
2, 9, 1 , 2 , 6
8
9
162.6 s
48.6 t
1ꢀ, 2ꢀ, 6ꢀ
3.92 (1H, d, J = 15.0)
3.17 (1H, d, J = 15.0)
8
2, 8
______
*Mutiplicities and coupling constants are in parentheses; assignment based on HMBC and HMQC experiments.
13
The C NMR and DEPT spectra revealed the presence of 14 signals attributed to a diketopiperazine unit at ꢁ 167.0 (s, C-2),
C
58.3 (d, C-3), 28.4 (t, C-4), 22.2 (t, C-5), 44.6 (t, C-6), 162.6 (s, C-8), and 48.6 (t, C-9) which was composed of glycine and
proline and a 2,3,6-tribromo-4,5-dihydroxylbenzyl unit at 124.3 (s, C-1ꢀ), 117.5 (s, C-2ꢀ), 114.4 (s, C-3ꢀ), 144.2 (s, C-4ꢀ), 146.9
1
1
(s, C-5ꢀ), 114.2 (s, C-6ꢀ), and 50.1 (t, C-7ꢀ) [4]. The diketopiperazine unit was confirmed by the H– H COSY between H-3 and
H -4, H -4 and H -5, and H -5 and H -6 and the HMBC correlations from H-3 to C-4 and C-7, H-4b to C-2 and C-5, and
2
2
2
2
2
H-9b to C-2 and C-8. The 2,3,6-tribromo-4,5-dihydroxylbenzyl unit was confirmed by detailed comparison of the above NMR
data (Experimental section) with that of 1 in the literature [7] and the HMBC correlations from H-7ꢀa to 1ꢀ, 2ꢀ, and 6ꢀ. The C-N
bond was confirmed by the HMBC correlations from H -7ꢀ to 1ꢀ, 2ꢀ and 6ꢀ. Compound 1 was established as 2-(2,3,6-tribromo-
2
4,5-dihydroxybenzyl)hexahydropyrrolo[1,2-a]pyrazine-1,4-dione.
The configuration of proline was further confirmed by HPLC analyses of the acid hydrolysate of 1 after derivatization
with the Marfey reagent (FDAA), allowing the absolute configurations at the ꢂ-carbon to be assigned as the L-configuration
for proline [8].
EXPERIMENTAL
IR spectra were recorded by the FT-IR microscope transmission method on a Nicolet 5700 FT-IR spectrophotometer.
1
13
NMR spectra were recorded on a Varian Inova 500 MHz spectrometer at 500.103 MHz for H and 125.762 MHz for C in
DMSO-d using solvent signals (DMSO, ꢁ 2.50/ꢁ 39.5) as reference; the coupling constants are in Hz. HR-ESI-MS data
6
H
C
were measured with an APEX II FT-ICR-MS spectrometer (Bruker Daltonics, Inc. USA). Column chromatography was
performed with silica gel (200–300 mesh, Qingdao Marine Chemical Inc. China) and Sephadex LH-20 (Pharmacia Biotech AB,
Uppsala, Sweden). HPLC separation was performed on an Agilent 1100 series and an Alltima 250 cm ꢃ 2.2 cm i.d. preparative
column packed with C18 (10 ꢄm). TLC was carried out with glass precoated silica gel GF254 plates. Spots were visualized
under UV light and by spraying with 2% FeCl in 95% EtOH.
3
Plant Material. Symphyocladia latiuscula was collected on the coast of Qingdao, Shandong Province, China in May
2004. The specimen was identified by Dr. Kui-Shuang Shao (Institute of Oceanology, ChineseAcademy of Sciences, Qingdao,
266071, China). A voucher specimen (No. 2004X16) was deposited at the Herbarium of the Institute of Oceanology, Chinese
Academy of Sciences, Qingdao, 266071, China.
Extraction and Isolation. The air-dried red alga Symphyocladia latiuscula (2.3 kg) was extracted with 95% EtOH at
room temperature for 3 ꢃ 72 h. After the solvent was removed under reduced pressure at < 40ꢅC, a dark residue (60 g) was
obtained. The residue was suspended in water and then partitioned with EtOAc. The EtOAc-soluble fraction (12 g) was
chromatographed over silica gel, eluting with a gradient of increasing Me CO (0–100%) in petroleum ether. The fraction
2
eluted by 30% Me CO in petroleum ether was rechromatographed over Sephadex LH-20 using petroleum ether–CHCl –MeOH
2
3
(5:5:1) to afford 18 subfractions. The ninth subfraction was further separated by reversed-phase preparative HPLC using
MeOH–H O–AcOH (75:20:0.1) as mobile phase to yield compound 1 (2.1 mg).
2
Absolute Configuration of Amino Acids [8]. Two set of compound 1 (50 ꢄg) were dissolved in 6 N HCl (400 ꢄL)
and put in 2 mL glass vials and heated at 110ꢅC for 24 h. Then the solvent was dried over nitrogen. The hydrolysate were
623

dissolved in 50 ꢄL distilled water and treated with a 50 mmol/L solution of FDAA (50 ꢄL) in acetone, followed by 1.0 N
NaHCO (20 ꢄL). The reaction mixtures were heated at 40ꢅC for 1.0 h, cooled to room temperature, and then acidified with
3
1.0 N HCl (20 ꢄL). In a similar fashion, standard L-proline and D-proline were derivatized separately. The derivatives of the
acid hydrolysate and the standard amino acids were subjected to Agilent reversed phase C18 column analysis (5 ꢄm, 4.6 ꢃ 150 mm;
1.0 mL/min; UV detection at 340 nm) with a linear gradient of (A) water–0.1% formic acid and (B) CH CN– 0.1% formic acid
3
from (35– 50%) over 15 min. The retention times for the FDAA derivatives of L-Pro and D-Pro were 8.3 and 9.2 min,
respectively.
2-(2,3,6-Tribromo-4,5-dihydroxybenzyl)-hexahydropyrrolo[1,2-a]pyrazine-1,4-dione (1). Brown amorphous
25
–1
powder; [ꢂ] –75ꢅ (c 0.1, MeOH). IR (
, cm ): 3247, 1718, 1660, 1554, 1450, 1399, 1289, 1179, 1406, 1024, 9879, 823,
D
max
+
79
765, 707. ESI-MS m/z: 511/513/515/516.8 (1:3:3:1) [M + H] ; HR-ESI-MS m/z 510.8509 (calcd for C H
510.8504). For H NMR and C NMR data, see Table 1.
Br N O ,
3 2 4
14 13
1
13
ACKNOWLEDGMENT
This work was supported by the Fundamental Research Funds for the Central Universities, Research Fund for the
Doctoral Program of Higher Education of China (200804911002), Scientific Fund of China University of Geosciences
(51900961135), National Natural Science Foundation of China (30901849), and Science and Technology Planning Project of
Zhejiang Province, China (2009C33004).
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